Acyl derivatives of vitamin b6



Patented Dec. 10, 1946 ACYL DERIVATIVES OF VITAMIN B6 John C. Keresztesy, Westfield, and Joseph R. Stevens, Rahway, N. J assignors to Merck & 00., Inc., Rahway, N. J., a corporation of New J ersey No Drawing. Original application April 30, 1940,

Serial No. 332,452. Divided and this application December 11, 1942, Serial No. 468,688

The present invention relates to a physiologically active chemical compound derived from vitamin Be and heretofore unknown as a chemical entity. This application is a division of our copending application Serial No. 332,452, filed April 30, 1940, which in turn is a division of our earlier application Serial No. 183,138, filed January 3,

1938, now matured into Patent No. sued June 23, 1942.

We have discovered that certain physiologically active chemical compounds can be obtained by treating materials containing the water soluble vitamin B6, either in free or in combined state as it is found naturally, which possess the unexhaving the physiological properties of vitamin B6 and a fat-soluble vitamin as, for instance, vitamins A or D in the same solvent. Another valuable inherent property of these oil soluble vitamin B6 derivatives is that they can be readily purified by sublimation in vacuo.

We have found that a suitable raw material for use in preparing these oil-soluble derivatives of vitamin B6 can be obtained by extracting rice bran with acidulated water, adsorbing the active principle on fullers earth, eluting and treating the eluate with an acylating agent. Of course, it is possible to prepare a suitable vitamin B6 concentrate by other methods obvious to those skilled in the art, or pure synthetic vitamin B6 itself can be used.

In preparing these new substances, an initial step can consist of treating an appropriate concentrate, of the kind heretofore described, with acid and with alkali to cause removal from, and chemical modification of, some of the inactive or foreign materials present. By inactive or foreign material is meant substances which are inactive per se or which possess a different physiological activity from vitamin B6, such as vitamin B1, nicotinic acid, lactoflavin, and other principles.

A modified procedure can include the removal of the inactive and/or chemically altered materials by extraction with appropriate solvents and subsequent separation of our physiologically active materials from these other inactive materials.

The final step consists of acylation of the vitamin-containing material and the subsequent separation of the desired acylated vitamin principle from foreign and other inactive or altered material, for example, by extraction with chloroform.

4 Claims. (01. 260-4295) If the active principle is to be isolated from extracts derived from natural products other than rice bran, the steps above outlined can be modified or supplemented by the precipitation of inactive material with ethyl nitrite and/or by a destruction by oxidation with mild oxidizing agents such as 10% nitric acid.

'By way of more specific illustration of the methods for producing the new substances, certain procedures of the type preferred above broadly outlined, will now be described.

A. Elimination of inactive or foreign materials by acid A We have found that a large amount of inactive materialcan be removed by means of an acid treatment, because insoluble compounds are formed which can be filtered off. and/or simple hydrolysis products are produced, which differ from their parent substances in their solubility, so that they can be removed by means of selective solvents in subsequent steps. Examples of preferred methods of practicing this step are presented by way of illustration:

About 500 grams of a vitamin-containing concentrate are dissolved in water to form about two liters of solution and 800 cc. of concentrated hydrochloric acid are added. This mixture (containing approximately 10% HCl) is allowed to digest on a steam bath for sixteen hours, then is cooled and the insoluble material removed by filtration. The solution is neutralized with 30% sodium hydroxide and the resulting mixture concentrated under vacuum to a syrupy mass which is then exhaustively extracted with ethyl alcohol.

B. Elimination of inactive and foreign material by alkali We have found also that many undesirable substances present in the concentrate can be removed by alkaline treatment without significant loss of vitamin B6. According to this method of treatment, the product obtained from process A is diluted with water and sufficient 30% sodium hydroxide is added to produce a solution con- Iacetone.

taining 10% sodium hydroxide. This solution is then digested on a steam bath for about sixteen hours at 100 C. then is cooled and filtered, neutralized with hydrochloric acid and evaporatedin vacuo to a syrup. This syrup is exhaustively extracted with 95% alcohol and the extract evaporated to a syrup free of alcohol.

fats, and fat solvents. The vitamin Be, as well as most of the impurities, are sparingly soluble in chloroform. We have found, however, that acylated vitamin B6 is easily extractable from aqueous solution with chloroform even in the presence of dilute acid. This difierencein solubility makes.

. it possible to remove a large amount of foreign By this treatment some inert materials ar re-' moved and also other inactive materials converted j into substances whose properties difier sufficient.-

1y from those of parent substances, so that they may b more easily removed in subsequent proc- I esses. In place of sodium hydroxide, other alkalis can be used, such as potassium, calcium, barium, or strontium, hydroxides,

C. Selective separation by meahs of solvents We have found that an appreciable concentration of the vitaminic substance can be effected by the use of certainsolvents. According to this procedure the; syrup obtained from step B is further concentrated to a thick gum in vacuo until it is practically free of water. This residue is then thoroughly extracted with hot absolute alcohol and the alcoholic solution, on cooling deposits insoluble inactive material which is removed by filtration. The alcoholic extract is then evaporated'to dryness on sand under high vacuum with rnild heating. The dried preparation is then subjected to a; series of exhaustive solvent extractions With several volumes of the solvent until further extractions do not remove significant quantities of soluble materials. The solvents used are ethyl ether, chloroform and acetone, prefer- Biological assay of the resulting fractions indicates that the vitamin hasbeen largely concentrated into the ably used successively in this order.

acetone extract with a substantial reduction of the. inert or inactive organic materials originally present.v In place of sand it is possible to use other adsorbent materials, such as powdered glass, asbestos and other inactive materials,- which will not form an adsorption complex with the activeprinciple, while permitting the distribution of the material to be extracted on'a large surface.

: The acetone extract can be further purified by redrying on sand under high vacuum, as before tice of the invention as disclosed, without 'ing from the spirit andjscope thereof.

mentioned, and again subjecting the material to repeated extractions with ether, cholorform and D. "Preparation of acylated vitamin LWehave found that under special conditions vitamin B6 can be converted to an acylated form in good yield which possesses the physiological "characteristics of the vitamin and which, as men- 'tioned, is valuable because of its solubility in oils,

bodies. ter-free extract obtained under C, containing 1 They product thusfobtained' acyl radicles of the aliphatic acids.

Our preferred method is to mix the wasmall amounts of alcohol, with acetyl chloride at room temperature, refluxing gently and, after cooling, treating. the reaction mixture with ice Water, extracting the acylated vitamin with chloroform, andrecovering the product by evaporating the chloroform. In this process acetyl chloride can bereplaced by acetic anhydride or by the chlorides or anhydrides of other lower aliphatic organic acids. The method of acylation will depend upon theacylating agent, solvent, etc.; such variations in procedur are well ,known to thoseskille'd in the, art. therapeutically pure and can be useddirectly'or jfurtherpurified if desired for special purposesj The examples ofuthev several stepsas herein set forth arepresented by way-of illustration to 'enablethose skilled-in the art to practice the same by appropriate adaptation to various source materials, and the invention is not intended to be limited to specific steps'so exemplified.

Obviously the number and order of, the steps, and solvents and materials, employed, may be modified, within reasonable limits, in the pracdepart- What is claimed is: 1 Fat soluble vitamin Bspreparations essentially consisting of therapeutically pure acetyl derivatives of vitamin B6 concentrates,

2. Fat-soluble vitamin B6 preparations essentially, consisting of therapeutically, pure acetyl derivatives of vitamin B6 in substantially pure form.

being the acylradicles of the lower molecular JoHN c. KEn szTEsvf JOSEPH R. s'rnvnns. 

